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Based on the surface plasmon resonance imaging (SPRi) instrument, we established a new method of analyzing morphine in urine by processing a calibration curve. According to an inhibition immunoassay, gradient concentration of morphine and morphine-BSA fixed on the chip competitively combine with morphine antibody on the chip. Given the three mathematical models, the data of SPRi signals generated from SPRi with morphine was processed to obtain the calibration curve. Ultrafiltration was used to pretreat blank urine samples with adding morphine, and then investigated the advantages and disadvantages of each model. With a limit detection of 6.57 ng·mL−1, the method and mathematical models can provide robust support for SPRi sensors used in further environmental detection, such as the epidemiological study of sewage.